Liquid chromatography–electrospray ionisation-mass spectrometry based method for the determination of estradiol benzoate in hair of cattle Original Research Article

A liquid chromatography (LC)-based method with mass spectrometric (MS/MS) detection was developed for the determination of estradiol benzoate residues in hair of cattle. First hair samples were pulverized with a ball mill followed by treatment with the reducing agent Tris(2-carboxyethyl)phosphine hydrochloride (TCEP). After liquid/liquid extraction samples were further purified by solid-phase extraction. Finally samples were analyzed with LC–MS/MS using deuterated estradiol benzoate as internal standard. The method was validated following the latest EU guidelines for screening methods using blank hair samples spiked at 5 ng g−1. The detection capability (CCβ) was less than 5 ng g−1 and the decision limit (CCα) was 1.6 ng g−1. The recovery was 27% at the 5 ng g−1 level and the accuracy was 95%. Confirmation, with two MS/MS transitions, was possible at a concentration of 5 ng g−1 or higher. Incurred samples obtained from different animal experiments were analyzed and the presence of estradiol benzoate was confirmed. Finally hair samples from different slaughterhouses were analyzed.